Genomic instability and tumor progression : a cytochemical, molecular biological and cytogenetic study of human tissue from uterine cervix, colon, breast and ovary

Detta är en avhandling från Stockholm : Karolinska Institutet, Department of Oncology-Pathology

Sammanfattning: Disruption of cell cycle checkpoint control is believed to be common in human carcinomas. To elucidate how cell cycle defects and genomic instability cooperate we investigated the pathogenesis of carcinomas by using four different models of tumor progression in human tissue samples i.e. from the uterine cervix, colon, breast and ovary. We describe how cell cycle proteins with different functions (Ki-67, cyclin A, cyclin E, p27KIP1, p53 and p21WAF1) are associated with different aspects of genomic instability in different stages of the progression from normal cells to the invasive malignant phenotype. In cervical tumorigenesis we found an increase in proliferative activity and tetraploidisation already in the mild dysplastic lesions. In severe dysplasia/CIS one case (1/10) had a gain of chromosome 3q. In the invasive carcinomas of early stage 90% (9/10) of the cases had a recurrent gain of 3q and in advanced stage tumors this was found in 77% of the cases, often with high-level copy number alterations. This result was closely related to aneuploidy and HPV 16 infection. However, recurrent gains and losses were mapped to several other chromosomal regions. Subchromosomal amplifications were mapped to chromosome arms 8q, 3q, and 5p. The results indicate that p53 inactivation by HPV, rather than p53 mutation and gain of 3q represents pivotal alterations in the progression of cervical carcinoma. In colorectal adenomas we found crude aneuploidy and high proliferative activity as early events. The transition to invasive carcinoma was characterized by the emergence of multiple chromosomal aberrations. In adenocarcinomas recurrent gains were mapped to chromosomes 1, 13 and 20 and to chromosome arms 7p and 8q. Losses were mapped to chromosome 4 and chromosome arms 8p and 18q. Small breast cancers (T1b, <10 mm had a low proliferative activity (MIB-1, cyclin A) and frequently low levels of p53. T1c tumors (11-20 mm) were frequently aneuploid (75%), with increased proliferative activity and p53 expression. In aneuploid tumors increasing numbers of chromosomal aberrations and regional amplifications were found with increasing tumor size (T1b to T1c). This was not seen in diploid tumors. In patients with longterm (>10 years)- and short-term survival (<4 years) CGH analysis revealed ANCA (average number of copy alterations) values of 8.3 and of 12.6 respectively. Aneuploid tumors from the long-term survivors had an ANCA value similar to that found in diploid tumors from both groups of patients. In tumors from short-term survivors recurrent gains in 3q, 9p, 11p and 11q and loss of 17p were observed and a threefold increase in the number of regional amplifications was found as compared to the long-term survivors. Furthermore, Southern blot hybridization performed on 117 breast carcinomas revealed gene amplifications in 44% of aneuploid tumors and in only 6% of diploid tumors. In ovarian carcinomas elevated expression of cyclin A and/or E was observed in almost all invasive carcinomas but only rarely in borderline tumors and in none of the benign cystadenomas. Similarly, increased numbers of tumor cells expressing high levels of p53 together with undetectable p21 were frequent events in carcinomas. Both borderline tumors and carcinomas revealed a high number of chromosomal gains and losses with the highest number of aberrations occurring in advanced stage carcinomas. The most pronounced difference between the borderline tumors and the invasive carcinomas was the number of regional amplifications, which were found to occur 13 fold more frequently in the carcinomas. The most frequent chromosomal gains were mapped to chromosome 7 and chromosome arms 8q, 3q and 1q in early stage carcinomas and to chromosome arms 3q, 8q and 5p in advanced stage carcinomas. Low expression of p27 together with high expression of MIB I detected by immunohistochemistry was found to be a strong significant predictor of poor outcome in ovarian carcinomas. Our results indicate that cytogenetic alterations occur with accelerated frequency in aneuploid tumors, possibly with disturbed cell cycle checkpoint control. Aggressive, or advanced, tumor variants can be identified by their specific chromosomal abnormalities and pattern of cell cycle disturbances.

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